Project Summary/Abstract Streptococcus pneumoniae (pneumococcus) is an important human pathogen and the most common cause of bacterial meningitis, pneumonia and otitis media. The ability to take up and incorporate foreign DNA has allowed pneumococcus to resist attempts at clearance by providing a resource for genetic adaptation. Genome analysis has demonstrated an enormous amount of genetic diversity largely derived from recombination with DNA from other streptococci. The competence state in pneumococcus is regulated by a quorum sensing system controlled by the com system which drives the production of a large number of genes including those involved in DNA uptake and recombination. Competent pneumococci can lyse non-competent members of the population releasing their DNA. Released DNA is taken up by competent cells and can serve as a source of new genetic material. We have identified an additional competitive locus called blp that is co- regulated with competence. It has the potential to expand the target range for DNA release through the production of bacteriocins (pneumocins). The blp locus is controlled by a parallel quorum sensing system to that required for competence induction. Competence induces the blp locus by upregulating the genes involved in blp specific pheromone secretion. Because the blp bacteriocins and their immunity proteins encoded in the blp cluster are variable from strain to strain, they can promote lysis of a wide range of pneumococci and related streptococcal species independent of the competence state of the target. In surveying the pneumococcal population, we found that 70% of isolates have a non-functional blp transporter. These strains lack inhibitory activity and fail to compete in the mouse nasopharynx due to a lack of pheromone and/or pneumocin secretion. We recently found that these strains can still induce their blp locus by using the competence transporter for secretion of the blp pheromone. In these strains, bacteriocin production is limited exclusively to periods of competence. To better understand the advantages of the transporter sufficient and deficient populations in adaptation and survival we have proposed three Aims. Aim 1 focuses on understanding how peptide concentrations are affected by cellular and environmental conditions and how this differs in the two populations. Aim 2 examines the implications of transporter integrity on competition and adaptation in vivo. Aim 3 will investigate how cross talk functions in structured communities such as biofilms using confocal imaging. These studies will take into account community structure, DNA persistence, media flow rate and peptide diffusion. In addition, the impact of the two populations on adaptation under these conditions will be evaluated. The persistence of this pathogen despite ongoing attempts at clearance from both the host and the surrounding flora is a testament to the power of genomic diversity and adaptation. Understanding how the variably expressed pneumocins contribute to this diversity will allow us to better anticipate and perhaps block attempts at escape from control measures via genetic exchange.